Scalable, diastereoselective synthesis of trans-Cyclooctenes with favorable physiochemical properties
Date
2022
Authors
Journal Title
Journal ISSN
Volume Title
Publisher
University of Delaware
Abstract
My doctoral research has centered on the synthesis and design of new chemical tools for bioorthogonal applications. Chapter 1 is an introductory chapter that describes the literature surrounding the synthesis of trans-cyclooctenes. This chapter emphasizes on the modern development of flow photochemical methods for preparative synthesis of trans-cyclooctene derivatives and includes an overview of the applications of trans-cyclooctenes in realms of synthesis and bioorthogonal chemistry. ☐ trans-Cyclooctenes (TCOs) are essential partners in the fastest known bioorthogonal reactions, but synthetic methods have been limited by poor diastereoselectivity. Especially hard to access are hydrophilic TCOs with favorable physicochemical properties for live cell or in vivo experiments. Described in Chapter 2 is a new class of TCOs, “a-TCOs”, prepared in high yield by stereocontrolled 1,2-additions of nucleophiles to trans-cyclooct-4-enone (keto-TCO). Computational transition state models rationalize the diastereoselectivity of 1,2-additions to deliver a-TCO products, which were also shown to be more reactive than standard TCOs and less hydrophobic than even a trans-oxocene analogue. Illustrating the favorable physicochemical properties of a-TCOs, a fluorescent TAMRA derivative in live HeLa cells was shown to be cell-permeable through intracellular Diels–Alder chemistry and to wash out more rapidly than other TCOs. I also describe a number of applications where a-TCOs are being utilized which include as sulfenic acid modifying probes for proteomic profiling in cell lysate, in the catalytic activation of bioorthogonal chemistry with light (CABL) at the suborganelle level in live cells, and as 18F-tracers for PET imaging applications in vivo. Lastly, I introduce a new regioselective synthesis for functionalized keto-TCOs that could be used to make trifunctional TCOs. In all, this chapter outlines a new, diastereoselective platform for trans-cyclooctene synthesis that increases the synthetic accessibility as well as improves the physiochemical properties of diversely functionalized trans-cyclooctenes for usage in bioorthogonal applications. ☐ In Chapter 3, I describe the development of a high throughput photochemical flow method employing a custom flow reactor for the large-scale synthesis of trans-cyclooctene derivatives as well as an improved workup protocol for trans-cyclooct-4-enone after photoisomerization. This custom flow reactor implements FEP tubing as an irradiation vessel in place of the previously utilized quartz flask as well as a higher yielding singlet sensitizer, 3,5-bis(trifluoromethyl)benzoate. This new flow reactor is amendable to 2-3 g and 12 g reaction scales, producing up to 8.6 g of trans-cyclooct-4-enone after 9 hours of irradiation of the cis-isomer at a rate of 1 g/h, greatly improving on the previous setup that produced 150 mg/h. In all, this chapter demonstrates improvements to both the throughput and workup efficiency of the photochemical flow synthesis of trans-cyclooctenes. The custom flow reactor has decreased the overall time needed to produce multigram quantities of trans-cyclooctenes, and thus has increased TCO synthetic accessibility for bioorthogonal applications. ☐ In Chapter 4, I describe protein proximity labeling methods for live cell proteomic profiling applications and introduce a new photocatalytic proximity labeling method that generates reactive electrophiles from n-alkylated dipyridyldihydrotetrazines for labeling of thiol nucleophiles which was demonstrated on both small molecules and protein targets. I describe a synthesis for n-alkylated and n-arylated dihydrotetrazines as well as a scalable synthesis of a small molecule nucleophilic addition adduct that I used for structural elucidation of the product by 1H NMR and high-resolution mass spectrometry analysis. In the future, this photocatalytic proximity labeling method will be tested on an endogenous protein target for live cell proteomic profiling applications.
Description
Keywords
Bioorthogonal applications, Trans-cyclooctene derivatives, Regioselective synthesis